Previous studies have suggested that odontoblasts sense Gram-positive bacteria components through Toll-like receptor 2 (TLR2) and trigger dental pulp immunity by producing specific pro-inflammatory cytokines and chemokines. However, most of the determinants of the inflammatory odontoblast response to these pathogens remain unknown. Objectives: The aim of this study was to identify inflammation-related pathways activated in vivo in bacteria-challenged inflamed human dental pulps and in vitro in odontoblast-like cells activated by two Gram-positive bacteria-derived TLR2 specific agonists. Methods: Gene expression of cyclooxygenase-2 (COX2), the NADPH oxidase subunits NOX1, NOX2, NOX3, NOX4, NOX5, DUOX1 and DUOX2, nitric oxide synthases 1, 2 and 3 (NOS1, NOS2 and NOS3) and heme oxygenase-1 (HMOX-1) was assessed by real-time PCR in healthy and bacteria-challenged inflamed dental pulps and in odontoblast-like cells stimulated with lipoteichoic acid (LTA), a cell wall component, or Pam2CSK4, a synthetic diacylated lipopeptide. NOS2 protein was localised in pulp samples by immunohistochemistry. NOS2 (inducible NOS) production was determined in Pam2CSK4-stimulated cells by western-blot. NOS activity and NO production were measured with Griess colorimetric assay in cells and culture supernatants, respectively. Results: Gene expression of COX2, the NADPH oxidase subunits NOX1, NOX2 and NOX4, NOS1, NOS2, NOS3, and HMOX1 were significantly increased in inflamed pulps compared to healthy ones. NOS2 was the most up-regulated gene. NOS2 protein was immunolocalized in odontoblasts beneath caries lesions but not in healthy samples. LTA stimulation of human odontoblast-like cells induced a low but significant increase of COX2, NOX1, NOX2, NOX5, DUOX1, NOS1, NOS2, NOS3 gene expression. Pam2CSK4 induced a strong increase of the same genes and of HMOX1. NOS2 protein production, NOS activity and NO release were clearly increased by Pam2CSK4. Conclusion: These results suggest that odontoblasts might contribute to the inflammatory response triggered in the human dental pulp by Gram-positive bacteria entering dentin during the caries process.